Biology subjects

Lewin, A. S.

Publications and source records attributed to Lewin, A. S..

5 recordsLinked to original sources

bioRxiv2023

Suppressor of Cytokine Signaling 3 Derived Peptide as a Therapeutic for Inflammatory, and Oxidative Stress Induced Damage to the Retina

PurposeInflammation and oxidative stress are contributing factors to age-related macular degeneration (AMD) and other retinal diseases. We tested a cell penetrating peptide from the kinase inhibitory region of intracellular checkpoint inhibitor Suppressor of Cytokine Signaling 3 (R9-SOCS3-KIR) peptide for its ability to blunt the inflammatory or oxidative pathways leading to AMD. MethodsWe used Anaphylatoxin C5a to mimic the effect of activated complement, lipopolysaccharide (LPS) and TNF to stimulate inflammation, and paraquat to induce mitochondrial oxidative stress. We used a human RPE cell line (ARPE-19) as proliferating cells and a mouse macrophage cell line (J774A.1) to follow cell propagation by microscopy or cell titer assays. We evaluated inflammatory pathways by monitoring nuclear translocation of NF-{kappa}B p65 and MAP kinase p38, and we used qRT-PCR and Western blots to evaluate induction of inflammatory markers. In differentiated ARPE-19 monolayers, we evaluated the integrity of tight junction proteins by microscopy and measurement of transepithelial electrical resistance. We used intraperitoneal injection of sodium iodate to test the ability of R9-SOC3-KIR to prevent RPE and retinal injury as assessed by fundoscopy, Optical Coherence Tomography (OCT) and histology. ResultsR9-SOCS3-KIR treatment suppressed C5a-induced nuclear translocation of the NF-kB activation domain p65 in undifferentiated ARPE-19 cells. TNF-mediated damage to tight junction proteins in RPE and the loss of transepithelial electrical resistance were prevented in the presence of R9-SOCS3-KIR. R9-SOCS3-KIR prevented the increased expression of genes related to inflammation in response to C5a treatment. R9-SOCS3-KIR also blocked lipopolysaccharide (LPS) induction of cyclooxygenase and inflammatory markers including IL-6, MCP1, COX-1 and IL-1{beta}. R9-SOCS3-KIR prevented paraquat mediated cell death and enhanced the levels of antioxidant effectors. Daily eye drop instillation of R9-SOCS3-KIR protected against retinal injury caused by i.p. administration of sodium iodate. ConclusionR9-SOCS3-KIR blocks the induction of inflammatory signaling in cell culture and reduces retinal damage in a widely used model of RPE/retina oxidative injury. Since this peptide can be administered by corneal instillation, this treatment may offer a convenient way to slow the progression of ocular diseases arising from inflammation and chronic oxidative stress.

immunology↗

bioRxiv2023

Modulation of Retinal Inflammation Delays Degeneration in a Mouse Model of Geographic Atrophy

The advanced form of AMD, geographic atrophy, is associated with increased RPE oxidative stress and chronic inflammation. Here we evaluated the effects of delivering an anti-inflammatory viral gene by an AAV-vector in a mouse model of geographic atrophy. We measured changes in retinal function, structure, and morphology over nine months with electroretinography, optical coherence tomography, and fundoscopy, respectively. In addition, we used retinal tissue to quantify changes in markers of inflammation by multiplex ELISA, RT-qPCR, and immunofluorescence staining. Our AAV significantly delayed the loss of retinal function and structure and decreased retinal inflammation compared to the control AAV treatment. Our results suggest that modulating retinal inflammation could significantly slow the progression of geographic atrophy.

neuroscience↗

bioRxiv2021

Binocular Benefit Following Monocular Subretinal AAV Injection in a Mouse Model of Autosomal Dominant Retinitis Pigmentosa (adRP)

Autosomal dominant retinitis pigmentosa (adRP) is frequently caused by mutations in RHO, the gene for rhodopsin. In previous experiments in dogs with the T4R mutation in RHO, an AAV2/5 vector expressing both an shRNA directed to human and dog RHO mRNA and an shRNA-resistant human RHO cDNA (AAV-RHO820-shRNA820) prevented retinal degeneration for more than 8 months following injection. To confirm that this same vector could protect the retinas of a different species and bearing a different RHO mutation, we injected mice transgenic for human P23H RHO at postnatal day 30 in one eye. For nine months, we monitored their retinal structure using spectral- domain optical coherence tomography (SD-OCT) and retinal function using electroretinography (ERG). We compared these to P23H RHO transgenic mice injected with AAV-GFP. Though retinas continued to thin over time, compared to control injected eyes, AAV-RHO820-shRNA820 slowed the loss of photoreceptor cells and decreased ERG amplitudes in AAV-RHO820-shRNA820 eyes during the nine-month study period. Unexpectedly, we also observed preservation of retinal structure and function in the untreated contralateral eyes of AAV-RHO820-shRNA820 treated mice. PCR analysis and western blots provided evidence that a low amount of vector from injected eyes was present in uninjected eyes.

neuroscience↗

bioRxiv2019

A Cell Penetrating Peptide from Type I Interferon Protects the Retina in a Mouse Model of Autoimmune Uveitis

Experimental autoimmune uveitis (EAU) in rodents recapitulates many features of the disease in humans and has served as a useful tool for the development of therapeutics. A peptide from C-terminus of interferon 1, conjugated to palmitoyl-lysine for cell penetration, denoted as IFN-C, was tested for its anti-inflammatory properties in ARPE-19 cells, followed by testing in a mouse model of EAU. Treatment with IFN-C and evaluation by RT-qPCR showed the induction of anti-inflammatory cytokines and chemokine. Inflammatory markers induced by treatment with TNF were suppressed when IFN-C was simultaneously present. TNF- mediated induction of NF-kB and signaling by IL-17A were attenuated by IFN-C. Differentiated ARPE-19 cells were treated with TNF in the presence or absence IFN-C and analyzed by immmunhistochemistry. IFN-C protected against the disruption integrity of tight junction proteins. Similarly, loss of transepithelial resistance caused by TNF was prevented by IFN-C. B10.RIII mice were immunized with a peptide from interphotoreceptor binding protein (IRBP) and treated by gavage with IFN-C. Development of uveitis was monitored by histology, fundoscopy, SD-OCT, and ERG. Treatment with IFN-C prevented uveitis in mice immunized with the IRBP peptide. Splenocytes isolated from mice with ongoing EAU exhibited antigenspecific T cell proliferation that was inhibited in the presence of IFN-C. IFN-C peptide exhibits anti-inflammatory properties and protects mice against damage to retinal structure and function suggesting that it has therapeutic potential for the treatment of autoimmune uveitis.

immunology↗

bioRxiv2018

A Cell Penetrating Peptide from SOCS-1 Prevents Ocular Damage in Experimental Autoimmune Uveitis

We describe an immunosuppressive peptide corresponding to the kinase inhibitory region (KIR) of the intracellular checkpoint protein suppressor of cytokine signaling 1 (SOCS-1) that binds to the phospho-tyrosine containing regions of the tyrosine kinases JAK2 and TYK2 and the adaptor protein MAL, and thereby inhibits signaling downstream from these signaling mediators. The peptide, SOCS1-KIR, is thus capable of downregulating overactive JAK/STAT or NF-kB signaling in somatic cells, including those in many compartments of the eye. Attachment of poly-arginine to this peptide (R9-SOCS1-KIR) allows it to penetrate the plasma membrane in aqueous media. R9-SOCS1-KIR was tested in ARPE-19 cells and was found to attenuate mediators of inflammation by blocking the inflammatory effects of IFN{gamma}, TNF, or IL-17A. R9-SOCS1-KIR also protected against TNF or IL-17A mediated damage to the barrier properties of ARPE-19 cells, as evidenced by immunostaining with the tight junction protein, zona occludin 1 (ZO-1), and measurement of transepithelial electrical resistance (TEER). Experimental autoimmune uveitis (EAU) was generated in B10.RIII mice using a peptide of interphotoreceptor retinal binding protein (IRBP161-180) as immunogen. Topical administration of R9-SOCS1-KIR protected ocular structure and function as seen by fundoscopy, optical coherence tomography (OCT), and electroretinography (ERG). The ability R9-SOCS1-KIR to suppress ocular inflammation and preserve barrier properties of retinal pigment epithelium makes it a potential candidate for aqueous treatment of autoimmune uveitis.\n\nHighlightsO_LIpeptide corresponding to the kinase inhibitory region of SOCS-1 linked to poly-arginine (R9-SOCS1-KIR) and its inactive control peptide were chemically synthesized.\nC_LIO_LIR9-SOCS1-KIR attenuated pro-inflammatory effects of IFN{gamma}, TNF, and IL-17 in ARPE-19 cells, thus showing a simultaneous inhibition Th1 and Th17 cell functions.\nC_LIO_LIDamage to barrier properties of ARPE-19 cells caused by TNF or IL-17 was prevented in the presence of R9-SOCS1-KIR.\nC_LIO_LITopical administration of R9-SOCS1-KIR prevented ocular damage in a mouse model of experimental autoimmune uveitis.\nC_LI

immunology↗